Rational Cisplatin based cancer therapy is an affordable and effective standard therapy for several solid cancers, including lung, ovarian and head and neck cancers. However, the clinical use of cisplatin is routinely limited by the development of drug resistance and subsequent therapeutic failure. Therefore, methods of circumventing cisplatin resistance have the potential to increase therapeutic efficiency and dramatically increase overall survival. Cisplatin resistance can be mediated by alterations to the DNA damage response, where multiple components of the repair machinery have been described to be client proteins of HSP90. In the present study, we have investigated whether therapy with the novel HSP90 inhibitor onalespib can potentiate the efficacy of cisplatin and potentially reverse cisplatin resistance in ovarian and head and neck cancer cells. Methods Cell viability, cancer cell proliferation and migration capacity were evaluatedin vitroon models of ovarian and head and neck cancer cells. Western blotting was used to assess the downregulation of HSP90 client proteins and alterations in downstream signaling proteins after exposure to cisplatin and/or onalespib. Induction of apoptosis and DNA damage response were evaluated in both monotherapy and combination therapy groups. Results Results demonstrate that onalespib enhances the efficiency of cisplatin in a dose-dependent manner. Tumor cells treated with both drugs displayed lower viability and a decreased migration rate compared to vehicle-control cells and cells treated with individual compounds. An increase of DNA double strand breaks was observed in both cisplatin and onalespib treated cells. The damage was highest and most persistent in the combination group, delaying the DNA repair machinery. Further, the cisplatin and onalespib co-treated cells had greater apoptotic activity compared to controls. Conclusion The results of this study demonstrate that the reduced therapeutic efficacy of cisplatin due to drug-resistance could be overcome by combination treatment with onalespib. We speculate that the increased apoptotic signaling, DNA damage as well as the downregulation of HSP90 client proteins are important mechanisms promoting increased sensitivity to cisplatin treatment.

Carcinoembryonic antigen (CEA) is an antigen that is highly expressed in colorectal cancers and widely used as a tumor marker. I-131 and Y-90-radiolabeled anti-CEA monoclonal antibodies (mAbs) have previously been assessed for radioimmunotherapy in early clinical trials with promising results. Moreover, the heat shock protein 90 inhibitor onalespib has previously demonstrated radiotherapy potentiation effects in vivo. In the present study, a Lu-177-radiolabeled anti-CEA hT84.66-M5A mAb (M5A) conjugate was developed and the potential therapeutic effects of Lu-177-DOTA-M5A and/or onalespib were investigated. The Lu-177 radiolabeling of M5A was first optimized and characterized. Binding specificity and affinity of the conjugate were then evaluated in a panel of gastrointestinal cancer cell lines. The effects on spheroid growth and cell viability, as well as molecular effects from treatments, were then assessed in several three-dimensional (3D) multicellular colorectal cancer spheroid models. Stable and reproducible radiolabeling was obtained, with labeling yields above 92%, and stability was retained at least 48 h post-radiolabeling. Antigen-specific binding of the radiolabeled conjugate was demonstrated on all CEA-positive cell lines. Dose-dependent therapeutic effects of both Lu-177-DOTA-M5A and onalespib were demonstrated in the spheroid models. Moreover, effects were potentiated in several dose combinations, where spheroid sizes and viabilities were significantly decreased compared to the corresponding monotherapies. For example, the combination treatment with 350 nM onalespib and 20 kBq Lu-177-DOTA-M5A resulted in 2.5 and 2.3 times smaller spheroids at the experimental endpoint than the corresponding monotreatments in the SNU1544 spheroid model. Synergistic effects were demonstrated in several of the more effective combinations. Molecular assessments validated the therapy results and displayed increased apoptosis in several combination treatments. In conclusion, the combination therapy of anti-CEA Lu-177-DOTA-M5A and onalespib showed enhanced therapeutic effects over the individual monotherapies for the potential treatment of colorectal cancer. Further in vitro and in vivo studies are warranted to confirm the current study findings.

Carcinoembryonic antigen (CEA) has emerged as an attractive target for theranostic applications in colorectal cancers (CRCs). In the present study, the humanized anti-CEA antibody hT84.66-M5A (M5A) was labeled with ¹⁷⁷Lu for potential CRC therapy. Moreover, the novel combination of ¹⁷⁷Lu-DOTA-M5A with the heat shock protein 90 inhibitor onalespib, suggested to mediate radiosensitizing properties, was assessed in vivo for the first time. M5A antibody uptake and therapeutic effects, alone or in combination with onalespib, were assessed in human CRC xenografts and visualized using SPECT/CT imaging. Although both ¹⁷⁷Lu-DOTA-M5A and onalespib monotherapies effectively reduced tumor growth rates, the combination therapy demonstrated the most substantial impact, achieving a fourfold reduction in tumor growth compared to the control group. Median survival increased by 33% compared to ¹⁷⁷Lu-DOTA-M5A alone, and tripled compared to control and onalespib groups. Importantly, combination therapy yielded comparable or superior effects to the double dose of ¹⁷⁷Lu-DOTA-M5A monotherapy. ¹⁷⁷Lu-DOTA-M5A increased apoptotic cell levels, indicating its potential to induce tumor cell death. These findings show promise for ¹⁷⁷Lu-DOTA-M5A as a CRC therapeutic agent, and its combination with onalespib could significantly enhance treatment efficacy. Further in vivo studies are warranted to validate these findings fully and explore the treatment’s potential for clinical use.

Simple summary

Cancer treatment is hampered by the limitations of individual therapy modalities and the intricate nature of the disease. The administration of maximal monotherapy doses often leads to undesirable side effects and/or therapy resistance. As a result, there is a growing recognition of the importance of investigating combination therapy to effectively address these obstacles. In the present in vivo study, the therapeutic effects of combination therapy with the heat shock protein 90 inhibitor onalespib, a potential radiosensitizer, and ¹⁷⁷Lu-DOTA-M5A for colorectal cancer (CRC) treatment were explored for the first time. The results demonstrated that the combination treatment was so effective that retained or even superior therapeutic effects could be achieved with only half the dose of administered ¹⁷⁷Lu-DOTA-M5A, showing enhanced tumor growth suppression and increased apoptosis. Consequently, the combination therapy involving ¹⁷⁷Lu-DOTA-M5A and onalespib constitutes a promising approach for treating metastatic CRCs. By enhancing therapeutic effects, minimizing therapy resistance, and reducing side effects, this approach has the potential to expand the patient population that can benefit from targeted treatment.

Radioimmunotherapy (RIT) is emerging as an effective treatment for metastatic solid tumors by coupling radionuclides with tumor-targeting molecules, precisely directing radiation to cancer cells while sparing healthy tissue. Carcinoembryonic antigen (CEA) is a promising target for RIT in CEA-expressing cancers, including colorectal cancers (CRCs). Recent studies highlight radiotherapy's role in enhancing the immune response against cancer. Combining RIT with immune checkpoint inhibitors, such as anti-PD-1 antibodies, may further enhance anti-tumor immunity and improve outcomes. This study aimed to investigate, for the first time, the in vivo effects of CEA-targeted RIT using the novel humanized anti-CEA antibody hT84.66-M5A labeled with ¹⁷⁷Lu (¹⁷⁷Lu-DOTA-M5A), combined with PD-1 blockade. Radioconjugate uptake and therapeutic effects were first assessed in vitro using the CRC spheroid model MC38-CEA1. The therapeutic effects of ¹⁷⁷Lu-DOTA-M5A and PD-1 blockade were then evaluated alone or in combination in CEA-transgenic mice bearing CEA-transduced CRC xenografts, with radioconjugate uptake validated in biodistribution studies and visualized via SPECT/CT imaging. Dose-dependent therapeutic effects of ¹⁷⁷Lu-DOTA-M5A were demonstrated in the 3D spheroid model. In vivo studies showed that both ¹⁷⁷Lu-DOTA-M5A and PD-1 antibody monotherapies effectively reduced tumor growth rates compared to the control group, but the combination therapy had the most significant impact. Combination therapy resulted in a dramatic tumor growth inhibition rate of -6% average daily, compared to +7%, +7.9%, and +13.5% in the PD-1 blockade, ¹⁷⁷Lu-DOTA-M5A (2.5 MBq), and control groups, respectively. Median survival increased by 31% in the PD-1 blockade group and by 52% in the ¹⁷⁷Lu-DOTA-M5A (2.5 MBq) group compared to the control group, while median survival was not reached in the corresponding combination group. Radioconjugate monotherapies and combination therapies did not introduce any bone marrow toxicity. ¹⁷⁷Lu-DOTA-M5A slightly altered the immune cell profile in the tumor microenvironment, increasing cytotoxic and helper T cells. Notably, pro-inflammatory macrophages became dominant over tumor-promoting ones in the tumor microenvironment of combination-treated mice. These findings highlight the promise of ¹⁷⁷Lu-DOTA-M5A as a CRC therapeutic agent and its enhanced efficacy when combined with an immune checkpoint inhibitor. Further in vivo studies are needed to fully validate these findings and explore the treatment’s potential for clinical use.